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- EMDB-5358: RNA-packaged Capsid of Bacteriophage phi6 -

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Basic information

Entry
Database: EMDB / ID: EMD-5358
TitleRNA-packaged Capsid of Bacteriophage phi6
Map dataRNA-packaged capsid of the bacteriophage phi6
Sample
  • Sample: RNA-packaged P1247 Capsid of Bacteriophage phi6
  • Virus: Bacteriophage phi6 (bacteriophage)
KeywordsdsRNA virus / Cystoviridae / capsid / expansion intermediate / segmented genome
Biological speciesBacteriophage phi6 (bacteriophage)
Methodsingle particle reconstruction / cryo EM / Resolution: 18.0 Å
AuthorsNemecek D / Cheng N / Qiao J / Mindich L / Steven AC / Heymann JB
CitationJournal: J Mol Biol / Year: 2011
Title: Stepwise expansion of the bacteriophage ϕ6 procapsid: possible packaging intermediates.
Authors: Daniel Nemecek / Naiqian Cheng / Jian Qiao / Leonard Mindich / Alasdair C Steven / J Bernard Heymann /
Abstract: The initial assembly product of bacteriophage ϕ6, the procapsid, undergoes major structural transformation during the sequential packaging of its three segments of single-stranded RNA. The ...The initial assembly product of bacteriophage ϕ6, the procapsid, undergoes major structural transformation during the sequential packaging of its three segments of single-stranded RNA. The procapsid, a compact icosahedrally symmetric particle with deeply recessed vertices, expands to the spherical mature capsid, increasing the volume available to accommodate the genome by 2.5-fold. It has been proposed that expansion and packaging are linked, with each stage in expansion presenting a binding site for a particular RNA segment. To investigate procapsid transformability, we induced expansion by acidification, heating, and elevated salt concentration. Cryo-electron microscopy reconstructions after all three treatments yielded the same partially expanded particle. Analysis by cryo-electron tomography showed that all vertices of a given capsid were either in a compact or an expanded state, indicating a highly cooperative transition. To benchmark the mature capsid, we analyzed filled (in vivo packaged) capsids. When these particles were induced to release their RNA, they reverted to the same intermediate state as expanded procapsids (intermediate 1) or to a second, further expanded state (intermediate 2). This partial reversibility of expansion suggests that the mature spherical capsid conformation is obtained only when sufficient outward pressure is exerted by packaged RNA. The observation of two intermediates is consistent with the proposed three-step packaging process. The model is further supported by the observation that a mutant capable of packaging the second RNA segment without previously packaging the first segment has enhanced susceptibility for switching spontaneously from the procapsid to the first intermediate state.
History
DepositionNov 28, 2011-
Header (metadata) releaseMay 23, 2012-
Map releaseMay 23, 2012-
UpdateMay 23, 2012-
Current statusMay 23, 2012Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 1.7
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 1.7
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_5358_1.jpg

Downloads & links

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Map

FileDownload / File: emd_5358.map.gz / Format: CCP4 / Size: 65.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRNA-packaged capsid of the bacteriophage phi6
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
2.63 Å/pix.
x 260 pix.
= 683.259 Å		2.63 Å/pix.
x 260 pix.
= 683.259 Å		2.63 Å/pix.
x 260 pix.
= 683.259 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 2.62792 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 2.07 / Movie #1: 1.7
Minimum - Maximum-2.55950069 - 5.63628864
Average (Standard dev.)0.0 (±1.0)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-130-130-130
Dimensions260260260
Spacing260260260
CellA=B=C: 683.25916 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.62791923076922.62791923076922.6279192307692
M x/y/z260260260
origin x/y/z0.0000.0000.000
length x/y/z683.259683.259683.259
α/β/γ90.00090.00090.000
start NX/NY/NZ-62-62-62
NX/NY/NZ125125125
MAP C/R/S123
start NC/NR/NS-130-130-130
NC/NR/NS260260260
D min/max/mean-2.5605.6360.000

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Supplemental data

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Sample components

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Entire : RNA-packaged P1247 Capsid of Bacteriophage phi6

EntireName: RNA-packaged P1247 Capsid of Bacteriophage phi6
Components
  • Sample: RNA-packaged P1247 Capsid of Bacteriophage phi6
  • Virus: Bacteriophage phi6 (bacteriophage)

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Supramolecule #1000: RNA-packaged P1247 Capsid of Bacteriophage phi6

SupramoleculeName: RNA-packaged P1247 Capsid of Bacteriophage phi6 / type: sample / ID: 1000
Oligomeric state: Icosahedral shell of P1 dimers with P2, P4 and P7 subunits and packaged RNA
Number unique components: 5
Molecular weightTheoretical: 12.6 MDa

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Supramolecule #1: Bacteriophage phi6

SupramoleculeName: Bacteriophage phi6 / type: virus / ID: 1 / Name.synonym: Bacteriophage phi6 / Sci species name: Bacteriophage phi6 / Database: NCBI / Virus type: OTHER / Virus isolate: STRAIN / Virus enveloped: Yes / Virus empty: Yes / Syn species name: Bacteriophage phi6
Host (natural)Organism: Pseudomonas syringae (bacteria) / synonym: BACTERIA(EUBACTERIA)
Molecular weightTheoretical: 12.6 MDa
Virus shellShell ID: 1 / Name: P1 / Diameter: 440 Å / T number (triangulation number): 1

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration10 mg/mL
BufferpH: 8 / Details: 10 mM potassium phosphate, 1 mM MgCl2, 100 mM NaCl
GridDetails: Quantifoil 2/2/400 mesh grid
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 93 K / Instrument: LEICA KF80 / Details: Vitrification instrument: Reichert - Jung KF80

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Electron microscopy

MicroscopeFEI/PHILIPS CM200FEG
TemperatureAverage: 93 K
Alignment procedureLegacy - Astigmatism: objective lens astigmatism was corrected at 100,000 times magnification
DateSep 24, 2008
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: NIKON SUPER COOLSCAN 9000 / Digitization - Sampling interval: 6.35 µm / Number real images: 32 / Average electron dose: 15 e/Å2 / Bits/pixel: 16
Electron beamAcceleration voltage: 120 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 48327 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.0 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 50000
Sample stageSpecimen holder: Gatan 626 / Specimen holder model: GATAN LIQUID NITROGEN

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Image processing

CTF correctionDetails: phase flip particle images
Final reconstructionAlgorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 18.0 Å / Resolution method: FSC 0.33 CUT-OFF / Software - Name: Bsoft / Number images used: 2659
FSC plot (resolution estimation)

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